Methods of treating erectile dysfunction

ABSTRACT

An active agent for use in a method of, or for use in the preparation of a medicament for, treating or inhibiting the development of erectile dysfunction and/or incontinence following pelvic radiation treatment in a subject in need thereof, comprising administering said subject an active agent in a treatment effective amount, is described. The active agent has the general structure of Formula I or a pharmaceutically acceptable salt thereof.

RELATED APPLICATIONS

This application is a 35 U.S.C. § 371 national phase entry of PCTApplication PCT/US2014/053442, filed Aug. 29, 2014, and published inEnglish on Mar. 12, 2015 as International Publication No. WO2015/034778, and which claims the benefit of U.S. Provisional PatentApplication Ser. No. 61/872,991, filed Sep. 3, 2013, the disclosure ofeach of which is incorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION

Prostate cancer is among the most common forms of cancer in men, and isfrequently treated with radiation therapy (alone, or in combination withother forms of treatment such as chemotherapy and surgery).Unfortunately, common side effects of prostate cancer treatment includeincontinence and erectile dysfunction. Given the wide-spread prevalenceof prostate cancer, there is accordingly a need for new approaches totreating these side effects.

SUMMARY OF THE INVENTION

A method of treating erectile dysfunction and/or incontinence followingpelvic radiation treatment in a subject in need thereof (e.g., a subjectadministered radiation treatment for cancer) is provided herein. Themethod comprises administering the subject an active agent as describedherein in an amount effective to treat erectile dysfunction and/orincontinence in the subject following the pelvic radiation treatment.

A still further aspect of the present invention is an active agent asdescribed herein for use in a method of treatment as described herein,or for use in the preparation of a medicament for carrying out a methodof treatment as described herein.

The present invention is explained in greater detail in thespecification set forth below. The disclosures of all United Statespatents cited herein are incorporated herein by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Diagram of Experimental Design. Rats received 5 consecutiveradiation doses of 7.5 Gy to the lower abdomen. Rats were injected i.p.with MnTE-2-PyP or PBS as a control throughout the study as indicated.Animals were harvested 12 weeks post-irradiation. There were fouranimals per group and the experiment was repeated once.

FIG. 2. Pharmacokinetics of MnTE-2-PyP in the rat urogenital system.Rats received injections of MnTE-2-PyP as outlined in FIG. 1 andharvested 1 week, 2 weeks, 12 weeks and 12 weeks+1 day after the startof injections. Rats harvested at 1 and 2 weeks had not received aninjection for 3 days (3 day trough). Rats harvested at 12 weeks, had notreceived an injection for 7 days (representing the lowest levels duringthe week) and 12 weeks+1 day represent rats 1 day after an injection(representing the highest levels during the week). A. Liver. B. Bowel.C. Prostate. D. Penile tissues. E. Bladder. MnTE-2-PyP was detected inthe liver, bowel, prostate, penile tissue and bladder. Data representsthe mean±standard error of the mean, n=3 per time point.

FIG. 3. Irradiated rats receiving MnTE-2-PyP lost significantly lessweight than irradiated rats injected with PBS. Weights of irradiatedanimals throughout the course of the experiment as compared to theirrespective non-irradiated groups. n=8 rats per group, asterisk (*)denotes p<0.05.

FIG. 4. Measuring erectile function in rats. Intracavernous pressure(ICP) was obtained after cavernous nerve stimulation as a measurement oferectile function 12 weeks post-irradiation. A. Representative pressurecurves obtained after nerve stimulation. B. Maximum ICP obtained aftercavernous nerve stimulation. Irradiation caused a significant decreasein ICP (RAD group) as compared to the non-irradiated group (PBS).MnTE-2-PyP significantly protected from the irradiation-induced loss inICP (MnTE-2-PyP RAD). n=8 rats/group, asterisk (*) denotes significantdifference from PBS group, p<0.05 and the number symbol (#) denotessignificant difference from RAD group, p<0.05.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

The present invention is primarily concerned with the treatment of humansubjects, but the invention may also be carried out on animal subjects,particularly mammalian subjects such as dogs, cats, livestock and horsesfor veterinary purposes. While subjects may be of any suitable age, thesubjects are in some embodiments neonatal, infant, juvenile, adolescent,adult, or geriatric subjects.

“Treat” as used herein refers to any type of treatment that imparts abenefit to a patient or subject matter as described herein, particularlydelaying or retarding the onset or progression of the conditionsdescribed herein, or reducing the severity of symptoms, or speeding orimproving recovery therefrom.

“Pharmaceutically acceptable” as used herein means that the compound orcomposition is suitable for administration to a subject to achieve thetreatments described herein, without unduly deleterious side effects inlight of the severity of the disease and necessity of the treatment.

“Combination” as used herein with respect to a method of administration(e.g., an active compound and an antibiotic administered in combination)includes administering the the two or more compounds simultaneously, orsequentially, sufficiently close in time to produce a combinedtherapeutic or treatment effect.

“Alkyl” as used herein alone or as part of another group, refers to astraight or branched chain hydrocarbon containing from 1 to 10 carbonatoms. Representative examples of alkyl include, but are not limited to,methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl,tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl,2,2-dimethylpentyl, 2,3-dimethylpentyl, n-heptyl, n-octyl, n-nonyl,n-decyl, and the like. “Lower alkyl” as used herein, is a subset ofalkyl, in some embodiments preferred, and refers to a straight orbranched chain hydrocarbon group containing from 1 to 4 carbon atoms.Representative examples of lower alkyl include, but are not limited to,methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, tert-butyl, andthe like. The term “akyl” or “loweralkyl” is intended to include bothsubstituted and unsubstituted alkyl or loweralkyl unless otherwiseindicated and these groups may be substituted with groups selected fromhalo (e.g., haloalkyl), alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl,cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl,hydroxyl, alkoxy (thereby creating a polyalkoxy such as polyethyleneglycol), alkenyloxy, alkynyloxy, haloalkoxy, cycloalkoxy,cycloalkylalkyloxy, aryloxy, arylalkyloxy, heterocyclooxy,heterocyclolalkyloxy, mercapto, alkyl-S(O)_(m), haloalkyl-S(O)_(m),alkenyl-S(O)_(m), alkynyl-S(O)_(m), cycloalkyl-S(O)_(m),cycloalkylalkyl-S(O)_(m), aryl-S(O)_(m), arylalkyl-S(O)_(m),heterocyclo-S(O)_(m), heterocycloalkyl-S(O)_(m), amino, carboxy,alkylamino, alkenylamino, alkynylamino, haloalkylamino, cycloalkylamino,cycloalkylalkylamino, arylamino, arylalkylamino, heterocycloamino,heterocycloalkylamino, disubstituted-amino, acylamino, acyloxy, ester,amide, sulfonamide, urea, alkoxyacylamino, aminoacyloxy, nitro or cyanowhere m=0, 1, 2 or 3.

“Alkenyl” as used herein alone or as part of another group, refers to astraight or branched chain hydrocarbon containing from 1 to 10 carbonatoms (or in loweralkenyl 1 to 4 carbon atoms) which include 1 to 4double bonds in the normal chain. Representative examples of alkenylinclude, but are not limited to, vinyl, 2-propenyl, 3-butenyl,2-butenyl, 4-pentenyl, 3-pentenyl, 2-hexenyl, 3-hexenyl, 2,4-heptadiene,and the like. The term “alkenyl” or “loweralkenyl” is intended toinclude both substituted and unsubstituted alkenyl or loweralkenylunless otherwise indicated and these groups may be substituted withgroups as described in connection with alkyl and loweralkyl above.

“Alkynyl” as used herein alone or as part of another group, refers to astraight or branched chain hydrocarbon containing from 1 to 10 carbonatoms (or in loweralkynyl 1 to 4 carbon atoms) which include 1 triplebond in the normal chain. Representative examples of alkynyl include,but are not limited to, 2-propynyl, 3-butynyl, 2-butynyl, 4-pentynyl,3-pentynyl, and the like. The term “alkynyl” or “loweralkynyl” isintended to include both substituted and unsubstituted alkynyl orloweralknynyl unless otherwise indicated and these groups may besubstituted with the same groups as set forth in connection with alkyland loweralkyl above.

“Cycloalkyl” as used herein alone or as part of another group, refers toa saturated or partially unsaturated cyclic hydrocarbon group containingfrom 3, 4 or 5 to 6, 7 or 8 carbons (which carbons may be replaced in aheterocyclic group as discussed below). Representative examples ofcycloalkyl include, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl, and cyclooctyl. These rings may be optionally substitutedwith additional substituents as described herein such as halo orloweralkyl. The term “cycloalkyl” is generic and intended to includeheterocyclic groups as discussed below unless specified otherwise.

“Heterocyclic group” or “heterocyclo” as used herein alone or as part ofanother group, refers to an aliphatic (e.g., fully or partiallysaturated heterocyclo) or aromatic (e.g., heteroaryl) monocyclic- or abicyclic-ring system. Monocyclic ring systems are exemplified by any 5or 6 membered ring containing 1, 2, 3, or 4 heteroatoms independentlyselected from oxygen, nitrogen and sulfur. The 5 membered ring has from0-2 double bonds and the 6 membered ring has from 0-3 double bonds.Representative examples of monocyclic ring systems include, but are notlimited to, azetidine, azepine, aziridine, diazepine, 1,3-dioxolane,dioxane, dithiane, furan, imidazole, imidazoline, imidazolidine,isothiazole, isothiazoline, isothiazolidine, isoxazole, isoxazoline,isoxazolidine, morpholine, oxadiazole, oxadiazoline, oxadiazolidine,oxazole, oxazoline, oxazolidine, piperazine, piperidine, pyran,pyrazine, pyrazole, pyrazoline, pyrazolidine, pyridine, pyrimidine,pyridazine, pyrrole, pyrroline, pyrrolidine, tetrahydrofuran,tetrahydrothiophene, tetrazine, tetrazole, thiadiazole, thiadiazoline,thiadiazolidine, thiazole, thiazoline, thiazolidine, thiophene,thiomorpholine, thiomorpholine sulfone, thiopyran, triazine, triazole,trithiane, and the like. Bicyclic ring systems are exemplified by any ofthe above monocyclic ring systems fused to an aryl group as definedherein, a cycloalkyl group as defined herein, or another monocyclic ringsystem as defined herein. Representative examples of bicyclic ringsystems include but are not limited to, for example, benzimidazole,benzothiazole, benzothiadiazole, benzothiophene, benzoxadiazole,benzoxazole, benzofuran, benzopyran, benzothiopyran, benzodioxine,1,3-benzodioxole, cinnoline, indazole, indole, indoline, indolizine,naphthyridine, isobenzofuran, isobenzothiophene, isoindole, isoindoline,isoquinoline, phthalazine, purine, pyranopyridine, quinoline,quinolizine, quinoxaline, quinazoline, tetrahydroisoquinoline,tetrahydroquinoline, thiopyranopyridine, and the like. These ringsinclude quaternized derivatives thereof and may be optionallysubstituted with groups selected from halo, alkyl, haloalkyl, alkenyl,alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo,heterocycloalkyl, hydroxyl, alkoxy, alkenyloxy, alkynyloxy, haloalkoxy,cycloalkoxy, cycloalkylalkyloxy, aryloxy, arylalkyloxy, heterocyclooxy,heterocyclolalkyloxy, mercapto, alkyl-S(O)_(m), haloalkyl-S(O)_(m),alkenyl-S(O)_(m), alkynyl-S(O)_(m), cycloalkyl-S(O)_(m),cycloalkylalkyl-S(O)_(m), aryl-S(O)_(m), arylalkyl-S(O)_(m),heterocyclo-S(O)_(m), heterocycloalkyl-S(O)_(m), amino, alkylamino,alkenylamino, alkynylamino, haloalkylamino, cycloalkylamino,cycloalkylalkylamino, arylamino, arylalkylamino, heterocycloamino,heterocycloalkylamino, disubstituted-amino, acylamino, acyloxy, ester,amide, sulfonamide, urea, alkoxyacylamino, aminoacyloxy, nitro or cyanowhere m=0, 1, 2 or 3.

“Aryl” as used herein alone or as part of another group, refers to amonocyclic carbocyclic ring system or a bicyclic carbocyclic fused ringsystem having one or more aromatic rings. Representative examples ofaryl include, azulenyl, indanyl, indenyl, naphthyl, phenyl,tetrahydronaphthyl, and the like. The term “aryl” is intended to includeboth substituted and unsubstituted aryl unless otherwise indicated andthese groups may be substituted with the same groups as set forth inconnection with alkyl and loweralkyl above.

“Arylalkyl” as used herein alone or as part of another group, refers toan aryl group, as defined herein, appended to the parent molecularmoiety through an alkyl group, as defined herein. Representativeexamples of arylalkyl include, but are not limited to, benzyl,2-phenylethyl, 3-phenylpropyl, 2-naphth-2-ylethyl, and the like.

“Heteroaryl” as used herein is as described in connection withheterocyclo above.

“Alkoxy” as used herein alone or as part of another group, refers to analkyl or loweralkyl group, as defined herein (and thus includingsubstituted versions such as polyalkoxy), appended to the parentmolecular moiety through an oxy group, —O—. Representative examples ofalkoxy include, but are not limited to, methoxy, ethoxy, propoxy,2-propoxy, butoxy, tert-butoxy, pentyloxy, hexyloxy and the like.

“Halo” as used herein refers to any suitable halogen, including —F, —Cl,—Br, and —I.

“Mercapto” as used herein refers to an —SH group.

“Azido” as used herein refers to an —N₃ group.

“Cyano” as used herein refers to a —CN group.

“Formyl” as used herein refers to a —C(O)H group.

“Carboxylic acid” as used herein refers to a —C(O)OH group.

“Hydroxyl” as used herein refers to an —OH group.

“Nitro” as used herein refers to an —NO₂ group.

“Acyl” as used herein alone or as part of another group refers to a—C(O)R radical, where R is any suitable substituent such as aryl, alkyl,alkenyl, alkynyl, cycloalkyl or other suitable substituent as describedherein.

“Alkylthio” as used herein alone or as part of another group, refers toan alkyl group, as defined herein, appended to the parent molecularmoiety through a thio moiety, as defined herein. Representative examplesof alkylthio include, but are not limited, methylthio, ethylthio,tert-butylthio, hexylthio, and the like.

“Amino” as used herein means the radical —NH₂.

“Alkylamino” as used herein alone or as part of another group means theradical —NHR, where R is an alkyl group.

“Arylalkylamino” as used herein alone or as part of another group meansthe radical —NHR, where R is an arylalkyl group.

“Disubstituted-amino” as used herein alone or as part of another groupmeans the radical —NR_(a)R_(b), where R_(a) and R_(b) are independentlyselected from the groups alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl,cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl.

“Acylamino” as used herein alone or as part of another group means theradical —NR_(a)R_(b), where R_(a) is an acyl group as defined herein andR_(b) is selected from the groups hydrogen, alkyl, haloalkyl, alkenyl,alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo,heterocycloalkyl.

“Acyloxy” as used herein alone or as part of another group means theradical —OR, where R is an acyl group as defined herein.

“Ester” as used herein alone or as part of another group refers to a—C(O)OR radical, where R is any suitable substituent such as alkyl,cycloalkyl, alkenyl, alkynyl or aryl.

“Amide” as used herein alone or as part of another group refers to a—C(O)NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitablesubstituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfoxyl” as used herein refers to a compound of the formula —S(O)R,where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl,alkynyl or aryl.

“Sulfonyl” as used herein refers to a compound of the formula —S(O)(O)R,where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl,alkynyl or aryl.

“Sulfonate” as used herein refers to a compound of the formula—S(O)(O)OR, where R is any suitable substituent such as alkyl,cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfonic acid” as used herein refers to a compound of the formula—S(O)(O)OH.

“Sulfonamide” as used herein alone or as part of another group refers toa —S(O)₂NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitablesubstituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Urea” as used herein alone or as part of another group refers to an—N(R_(c))C(O)NR_(a)R_(b) radical, where R_(a), R_(b) and R_(c) are anysuitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl oraryl.

“Alkoxyacylamino” as used herein alone or as part of another grouprefers to an —N(R_(a))C(O)OR_(b) radical, where R_(a), R_(b) are anysuitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl oraryl.

“Aminoacyloxy” as used herein alone or as part of another group refersto an —OC(O)NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitablesubstituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

1. Active Compounds/Active Agents.

Active compounds or active agents of the present invention are, ingeneral, porphyrin compounds. The active compounds include superoxidedismutase (SOD) mimetic prophryin compounds, and/or redox activeporphyrin compounds.

Examples of porphyrin active compounds, and methods of making the same,include but are not limited to those set forth in U.S. Pat. No.8,470,808 to Piganelli et al.; U.S. Pat. No. 8,183,364 toBatinic-Haberle et al., U.S. Pat. No. 6,916,799 to Fridovich et al.;U.S. Pat. No. 6,479,477 to Crapo et al.; U.S. Pat. No. 6,583,132 toCrapo et al. and in US Patent Application Pub. No. US 2012/0065181 toWarner et al.; the disclosures of which are incorporated by referenceherein in their entirety.

Examples of active compounds include but are not limited to compounds ofFormula I:

wherein:

each R is independently substituted or unsubstituted aryl, heteroaryl,cycloalkyl, or heterocycloalkyl;

each A is an independently selected hydrogen, or an electron-withdrawingor electron donating group (e.g., e.g., is halogen, NO₂ or CHO),

M is a metal, e.g., selected from the group consisting of manganese,iron, copper, cobalt, nickel and zinc, or is absent (in which case ahydrogen is added to each of the two nitrogens required to correctvalency), and

Z⁻is a counterion.

In some embodiments of Formula I above, each R is preferably heteroarylor heterocycloalkyl, particularly those containing at least one or twonitrogen atoms in the heterocyclic ring (e.g., pyrrolyl, imidazolyl,triazolyl, pyridyl, pyrimidyl, triazinyl, oxazolyl, thiazolyl, oxazinyl,thiazinyl, oxathiazinyl, etc.), in some embodiments wherein at least oneof which nitrogen atoms (or in some embodiments at least two of whichnitrogen atoms) are optionally but preferably substituted (e.g.,quaternized) with a substituent such as described in connection withheterocyclic groups above (e.g., substituted with alkyl, alkoxyalkyl,etc.).

Still more particular examples of the foregoing active compounds includebut are not limited to those set forth below.

A. Alkyl substituted imidazole porphyrins. In some embodiments theactive compound has a structure of Formula A1 or A2:

wherein:

each R is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straightchain or branched);

each A is, independently, hydrogen or an electron withdrawing group(e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron,copper, cobalt, nickel and zinc, and

Z⁻is a counterion.

In some embodiments the active compound has the formula:

wherein Z− is a counterion.

B. Alkyl substituted pyridyl porphyrins. In some embodiments the activecompound has a structure of Formula B1 or B2:

wherein:

each R is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straightchain or branched);

each A is, independently, hydrogen or an electron withdrawing group(e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron,copper, cobalt, nickel and zinc, and

Z⁻is a counterion.

In some embodiments the compound has a structure of the Formula V:

wherein each R, A, M and Z is as given in connection with Formula B1 andB2 above.

In some embodiments the compound has the structure:

wherein Z⁻is a counterion.

C. Alkoxyalkyl substituted pyridyl porphyrins. In some embodiments theactive compound has a structure of Formula C1 or C2:

wherein:

each R is —(CH₂)_(m)CH₂OX;

m is 1 or 2, preferably 1;

X is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straightchain or branched).

each A is, independently, hydrogen or an electron withdrawing group(e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron,copper, cobalt, nickel and zinc, and

Z⁻is a counterion.

In some embodiments the compound has a structure of the Formula V:

wherein each R, A, M and Z is as given in connection with Formula C1 andC2 above.

In some embodiments the compound has the structure:

wherein Z⁻is a counterion.

D. Salts. The active compounds disclosed herein can, as noted above, beprepared in the form of their salts or pharmaceutically acceptablesalts, e.g., to provide a compound or composition including a counterionas noted above. Pharmaceutically acceptable salts are salts that retainthe desired biological activity of the parent compound and do not impartundesired toxicological effects. Examples of such salts are (a) acidaddition salts formed with inorganic acids, for example hydrochloricacid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid andthe like; and salts formed with organic acids such as, for example,acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid,fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid,benzoic acid, tannic acid, palmitic acid, alginic acid, polyglutamicacid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonicacid, naphthalenedisulfonic acid, polygalacturonic acid, and the like;(b) salts formed from elemental anions such as chlorine, bromine, andiodine, and (c) salts derived from bases, such as ammonium salts, alkalimetal salts such as those of sodium and potassium, alkaline earth metalsalts such as those of calcium and magnesium, and salts with organicbases such as dicyclohexylamine and N-methyl-D-glucamine.

2. Pharmaceutical Formulations.

The active compounds described above may be formulated foradministration in a pharmaceutical carrier in accordance with knowntechniques. See, e.g., Remington, The Science And Practice of Pharmacy(9^(th) Ed. 1995). In the manufacture of a pharmaceutical formulationaccording to the invention, the active compound (including thephysiologically acceptable salts thereof) is typically admixed with,inter alia, an acceptable carrier. The carrier must, of course, beacceptable in the sense of being compatible with any other ingredientsin the formulation and must not be deleterious to the patient. Thecarrier may be a solid or a liquid, or both, and is preferablyformulated with the compound as a unit-dose formulation, for example, atablet, which may contain from 0.01 or 0.5% to 95% or 99% by weight ofthe active compound. One or more active compounds may be incorporated inthe formulations of the invention, which may be prepared by any of thewell known techniques of pharmacy comprising admixing the components,optionally including one or more accessory ingredients.

The formulations of the invention include those suitable for oral,rectal, topical, buccal (e.g., sub-lingual), vaginal, parenteral (e.g.,subcutaneous, intramuscular, intradermal, or intravenous), topical(i.e., both skin and mucosal surfaces, including airway surfaces) andtransdermal administration, although the most suitable route in anygiven case will depend on the nature and severity of the condition beingtreated and on the nature of the particular active compound which isbeing used.

Formulations suitable for oral administration may be presented indiscrete units, such as capsules, cachets, lozenges, or tablets, eachcontaining a predetermined amount of the active compound; as a powder orgranules; as a solution or a suspension in an aqueous or non-aqueousliquid; or as an oil-in-water or water-in-oil emulsion. Suchformulations may be prepared by any suitable method of pharmacy whichincludes the step of bringing into association the active compound and asuitable carrier (which may contain one or more accessory ingredients asnoted above). In general, the formulations of the invention are preparedby uniformly and intimately admixing the active compound with a liquidor finely divided solid carrier, or both, and then, if necessary,shaping the resulting mixture. For example, a tablet may be prepared bycompressing or molding a powder or granules containing the activecompound, optionally with one or more accessory ingredients. Compressedtablets may be prepared by compressing, in a suitable machine, thecompound in a free-flowing form, such as a powder or granules optionallymixed with a binder, lubricant, inert diluent, and/or surfaceactive/dispersing agent(s). Molded tablets may be made by molding, in asuitable machine, the powdered compound moistened with an inert liquidbinder.

Formulations suitable for buccal (sub-lingual) administration includelozenges comprising the active compound in a flavoured base, usuallysucrose and acacia or tragacanth; and pastilles comprising the compoundin an inert base such as gelatin and glycerin or sucrose and acacia.

Formulations of the present invention suitable for parenteraladministration comprise sterile aqueous and non-aqueous injectionsolutions of the active compound(s), which preparations are preferablyisotonic with the blood of the intended recipient. These preparationsmay contain anti-oxidants, buffers, bacteriostats and solutes whichrender the formulation isotonic with the blood of the intendedrecipient. Aqueous and non-aqueous sterile suspensions may includesuspending agents and thickening agents. The formulations may bepresented in unit\dose or multi-dose containers, for example sealedampoules and vials, and may be stored in a freeze-dried (lyophilized)condition requiring only the addition of the sterile liquid carrier, forexample, saline or water-for-injection immediately prior to use.Extemporaneous injection solutions and suspensions may be prepared fromsterile powders, granules and tablets of the kind previously described.For example, in one aspect of the present invention, there is providedan injectable, stable, sterile composition comprising an activecompound(s), or a salt thereof, in a unit dosage form in a sealedcontainer. The compound or salt is provided in the form of alyophilizate which is capable of being reconstituted with a suitablepharmaceutically acceptable carrier to form a liquid compositionsuitable for injection thereof into a subject. The unit dosage formtypically comprises from about 10 mg to about 10 grams of the compoundor salt. When the compound or salt is substantially water-insoluble, asufficient amount of emulsifying agent which is physiologicallyacceptable may be employed in sufficient quantity to emulsify thecompound or salt in an aqueous carrier. One such useful emulsifyingagent is phosphatidyl choline.

Formulations suitable for rectal administration are preferably presentedas unit dose suppositories. These may be prepared by admixing the activecompound with one or more conventional solid carriers, for example,cocoa butter, and then shaping the resulting mixture.

Formulations suitable for topical application to the skin preferablytake the form of an ointment, cream, lotion, paste, gel, spray, aerosol,or oil. Carriers which may be used include petroleum jelly, lanoline,polyethylene glycols, alcohols, transdermal enhancers, and combinationsof two or more thereof.

Formulations suitable for transdermal administration may be presented asdiscrete patches adapted to remain in intimate contact with theepidermis of the recipient for a prolonged period of time. Formulationssuitable for transdermal administration may also be delivered byiontophoresis (see, for example, Pharmaceutical Research 3 (6):318(1986)) and typically take the form of an optionally buffered aqueoussolution of the active compound. Suitable formulations comprise citrateor bis\tris buffer (pH 6) or ethanol/water and contain from 0.1 to 0.2Mactive ingredient.

Further, the present invention provides liposomal formulations of thecompounds disclosed herein and salts thereof. The technology for formingliposomal suspensions is well known in the art. When the compound orsalt thereof is an aqueous-soluble salt, using conventional liposometechnology, the same may be incorporated into lipid vesicles. In such aninstance, due to the water solubility of the compound or salt, thecompound or salt will be substantially entrained within the hydrophiliccenter or core of the liposomes. The lipid layer employed may be of anyconventional composition and may either contain cholesterol or may becholesterol-free. When the compound or salt of interest iswater-insoluble, again employing conventional liposome formationtechnology, the salt may be substantially entrained within thehydrophobic lipid bilayer which forms the structure of the liposome. Ineither instance, the liposomes which are produced may be reduced insize, as through the use of standard sonication and homogenizationtechniques.

Of course, the liposomal formulations containing the compounds disclosedherein or salts thereof, may be lyophilized to produce a lyophilizatewhich may be reconstituted with a pharmaceutically acceptable carrier,such as water, to regenerate a liposomal suspension.

Other pharmaceutical compositions may be prepared from thewater-insoluble compounds disclosed herein, or salts thereof, such asaqueous base emulsions. In such an instance, the composition willcontain a sufficient amount of pharmaceutically acceptable emulsifyingagent to emulsify the desired amount of the compound or salt thereof.Particularly useful emulsifying agents include phosphatidyl cholines,and lecithin.

In addition to active compound(s), the pharmaceutical compositions maycontain other additives, such as pH-adjusting additives. In particular,useful pH-adjusting agents include acids, such as hydrochloric acid,bases or buffers, such as sodium lactate, sodium acetate, sodiumphosphate, sodium citrate, sodium borate, or sodium gluconate. Further,the compositions may contain microbial preservatives. Useful microbialpreservatives include methylparaben, propylparaben, and benzyl alcohol.The microbial preservative is typically employed when the formulation isplaced in a vial designed for multidose use. Of course, as indicated,the pharmaceutical compositions of the present invention may belyophilized using techniques well known in the art.

3. Dosage and Routes of Administration.

As noted above, the present invention provides pharmaceuticalformulations comprising the active compounds (including thepharmaceutically acceptable salts thereof), in pharmaceuticallyacceptable carriers for oral, rectal, topical, buccal, parenteral,intramuscular, intradermal, or intravenous, and transdermaladministration.

The effective amount (e.g., therapeutically effective or treatmenteffective amount) or dosage of any specific active compound as describedherein, for use in any specific method as described herein, will varydepending on factors such as the condition being treated, the route ofadministration, the general condition of the subject (e.g., age, gender,weight, etc.), etc. In general (e.g., for oral or parenteraladministration), the dosage may be from about 0.01, 0.05, or 0.1milligram per kilogram subject body weight (mg/kg), up to about 1, 5, or10 mg/kg. For topical administration, the active agent may be includedin a pharmaceutically acceptable composition to be applied in anysuitable amount, typically from 0.01, 0.1, or 1 percent by weight, up to10, 20, or 40 percent by weight, or more, of the weight of thecomposition, again depending on factors such as the condition beingtreated, condition of the subject, etc.

The active agents described herein may be administered directly orthrough the administration to the subject of a pharmaceuticallyacceptable prodrug which is in turn converted to the active agent invivo. The term “prodrug” refers to compounds that are rapidlytransformed in vive to yield the parent compound of the above formulae,for example, by hydrolysis in blood. A thorough discussion is providedin T. Higuchi and V. Stella, Prodrugs as Novel delivery Systems, Vol. 14of the A.C.S. Symposium Series and in Edward B. Roche, ed.,Bioreversible Carriers in Drug Design, American PharmaceuticalAssociation and Pergamon Press, 1987, both of which are incorporated byreference herein. See also U.S. Pat. No. 6,680,299 Examples include aprodrug that is metabolized in vivo by a subject to an active drughaving an activity of active compounds as described herein, wherein theprodrug is an ester of an alcohol or carboxylic acid group, if such agroup is present in the compound; an acetal or ketal of an alcoholgroup, if such a group is present in the compound; an N-Mannich base oran imine of an amine group, if such a group is present in the compound;or a Schiff base, oxime, acetal, enol ester, oxazolidine, orthiazolidine of a carbonyl group, if such a group is present in thecompound, such as described in U.S. Pat. Nos. 6,680,324 and 6,680,322.

4. Treating Radiation-induced Erectile Dysfunction and/or Incontinence.

As noted above, the present invention provides a method of treatingerectile dysfunction and/or incontinence (including both urinary andbowel incontinence) following pelvic radiation or irradiation treatment.The subject may be a male subject who has received pelvic radiation orirradiation treatment for prostate cancer; a female subject who hasreceived pelvic radiation or irradiation treatment for cervical cancer,uterine cancer, or endometrial cancer; or a male or female subject whohas received pelvic radiation or irradiation treatment for other cancerslocated in the pelvic region (e.g., for colorectal cancer or bladdercancer).

The radiation treatment may comprise or have comprised external beamradiation, radioactive seed implantation therapy (brachytherapy), or acombination thereof.

The cancer may comprise or have comprised adenocarcinoma or small cellcarcinoma, and may comprise or have comprised Stage I, Stage IIA, StageIIB, Stage III, or Stage IV cancer.

The active agent may be administered on any suitable schedule, includingprior to the onset of radiation treatment, during radiation treatment,and/or following radiation treatment of the subject. The active agentmay be administered in the dosages described above on any suitableschedule, e.g., once, twice, or three times daily, for any suitableduration, typically from 1 or 2 weeks, up to 1 or 2 months or more,depending on the duration of the radiation treatment, and whenadministration of the active agent is initiated.

The present invention is explained in greater detail in the followingnon-limiting Examples.

Experimental 1 Manganese (III)Meso-Tetrakis-(N-Methylpyridinium-2-yl)porphyrin (MnTE-2-PyP) preventsside-effects Incurred by Pelvic Irradiation

To test whether MnTE-2-PyP would protect the urogenital system fromdamage associated with pelvic irradiation, rats were exposed tofractionated irradiation of the pelvic region with and withoutMnTE-2-PyP administration. Animals were harvested 12 weekspost-irradiation and it was found that MnTE-2-PyP protected thestructure and function of organs exposed to radiation. Specifically,MnTE-2-PyP protected the skin, prostate, testes, and penile tissues fromirradiation-induced damage and prevented the loss of erectile functioncaused by radiation therapy.

Materials and Method.

Experimental animals. Sprague-Dawley rats (100-150 g, 4-6 weeks in age)from either Jackson or Harlan Laboratories were used in this study. Ratswere housed at either the University of Colorado Anschutz Medical Campusor at National Jewish Health and given a continuous supply of food andwater. This study was carried out in strict accordance with therecommendations of the Guide for the Care and Use of Laboratory Animalsof the National Institutes of Health. All treatments and procedures wereapproved by the institutional animal care and use committees at the twoinstitutions.

Treatment groups and experimental design. Animals were randomly assignedto 4 groups (8 rats per group): 1) PBS injected with no irradiation, 2)MnTE-2-PyP injected with no irradiation, 3) PBS injected withirradiation, and 4) MnTE-2-PyP injected with irradiation. Theexperimental design is illustrated in FIG. 1. Briefly, animals wereinjected with MnTE-2-PyP (5 mg/kg) or PBS 24 hours before the start ofradiation. Rats were irradiated for 5 sequential days with 7.5 Gy/day inthe lower pelvic region. Animals were positioned under a lead shieldsuch that only the lower pelvic region received radiation. Radiation wasdelivered by X-ray irradiation at a dose rate of 1 Gy per minute. Thisirradiation scheme was chosen because it mimics an irradiation schemethat a patient undergoing prostate cancer therapy would undertake toeradicate the prostate tumor (G. van der Wielen et al., J Sex Med 6:1908-1913). MnTE-2-PyP (2.5 mg/kg) or PBS was administered every otherday for the following two weeks. The animals were then administeredMnTE-2-PyP (5 mg/kg) or PBS once a week until 12 weeks post-irradiation.The rats weights and radiation-induced skin damage was documented once aweek. At the 12 week time point, the erectile function of the rats wasaccessed and tissues (prostate and penis) were harvested from theanimals. This experiment was repeated once with the same groups and timepoints.

MnTE-2-PyP treatment. MnTE-2-PyP was synthesized by Ricerca Biosciences,LLC. Concord, Ohio, USA. MnTE-2-PyP was dissolved in PBS and injected(100 μl) intraperitoneally (i.p.) at 2.5 or 5 mg/kg at the abovespecified times. PBS (100 μl) injected i.p. was used as a control.

Pharmacokinetics of MnTE-2-PyP in rat urogenital system. The sameMnTE-2-PyP dosing scheme was used for pharmacokinetic analysis as wasused for the experimental design with a different cohort of animals.There were four groups of animals and each group had 3 rats: group 1)rats were harvested 1 week after the start of injections, group 2) ratswere harvested 2 weeks after the start of injections, group 3) rats wereharvested 12 weeks after the start of injections (harvested 7 days afterthe last injection), group 4) rats were harvested 12 weeks+ one dayafter the start of injections (harvested 1 day after the lastinjection). At each time point, the liver, bowel, prostate, peniletissue and bladder were collected and flash frozen. The MnTE-2-PyPconcentrations were then determined by the PK/PD Bioanalytical CoreLaboratory at the Duke Cancer Institute as previously described (I.Spasojevic et al., Free Radic Biol Med 45: 943-949).

Scoring of skin reaction to irradiation. Animals were observed throughthe course of the experiment. They received a score from a reactionscale from Hall and Giaccia [24]. O=no visible reaction, 1=fainterythema, 2=erythema, 3=marked erythema, 4=moist desquamation of lessthan half of the irradiated area, 5=moist desquamation of more than halfthe irradiated area.

Histology of rat urogenital system. Rat tissues (prostate and penis)were fixed in 4% paraformaldehyde and embedded in paraffin. The tissueblocks were sectioned (5 μm thick), deparaffinized in xylene andrehydrated through sequential steps of 100, 95, and 75% ethanol. Thesections were stained with hematoxylin and eosin, dehydrated andcoverslips mounted. Penile tissue was also stained with trichrome(Sigma, St. Louis, Mo.) to visualize fibrosis, according to themanufacturer's protocol.

Immunohistochemistry. Immunohistochemistry was performed utilizingstandard DAB techniques (Vector Labs, Burlingame, Calif.). Paraffinblocks were sectioned, deparaffinized, and rehydrated. Antigens wereunmasked with preheated antigen retrieval in sodium citrate buffer for20 min and then cooled. H₂O₂ (3%) was placed on the slides for 5 minutesto block endogenous peroxides, followed by a serum block for 30 minutes.The 8-OHdG primary antibody (Abcam, Cambridge, Mass., 5 μg/mL) was thenadded overnight at the indicated dilution. Biotinylated mouse secondaryantibody (1:200) was then added for 45 minutes. Biotin binding wasincreased using the ABC Elite method for 30 min and stains werevisualized with DAB. The tissues were counterstained with hematoxylin,dehydrated and coversliped. The 8-OHdG stained sections were quantifiedby taking 10 random pictures of each prostate section. The sections werescored blindly, and any nuclear staining was counted as a positive8-OHdG stained cell.

Erectile functional assay. The erectile functional assay was performedin rats 12 weeks post-irradiation, as previously described (J. Dall'Eraet al., Int J Impot Res 20: 307-314.). Briefly, rats were anesthetizedwith sodium pentobarbital before dissection of the lower abdomen tolocate the cavernous nerve. Two stainless steel electrodes were placedaround the cavernous nerve on one side, with the negative electrodeapproximately 1 mm from the positive electrode. The skin overlaying thepenis was removed and the crura were dissected free. A 26-gauge needlethat was connected to a pressure transducer and was inserted into eitherthe right or left crus. Electrostimulation was performed using astimulator (World Precision Instruments, Sarasota, Fla., USA). Increasesin intracavernous pressure were measured and recorded using theData-Trax data acquisition software (Distributed Design Concepts, Dover,N.H., USA).

Results

Pharmacokinetic analysis was performed to ensure that adequate levels ofMnTE-2-PyP would reach the urogenital tissues when administeredintraperitoneally (i.p.). Rats were subjected to the same MnTE-2-PyPdosing scheme (FIG. 1) as the animals undergoing irradiation and tissueswere harvested 1 week, 2 weeks, 12 weeks, and 12 weeks+ one day afterthe start of drug administration. The liver, bowel, prostate, penis, andbladder tissues all contained MnTE-2-PyP at every time pointinvestigated (FIG. 2). The liver contained the highest concentration ofMnTE-2-PyP, which is not surprising since MnTE-2-PyP is known toaccumulate in the liver (I. Spasojevic et al., Free Radic Biol Med 45:943-949). The bowel, prostate, and bladder all had similar levels ofMnTE-2-PyP (˜1000 nM) over the course of the pharmacokinetic analysis.This concentration of MnTE-2-PyP has been shown to prevent free radicaldamage in other tissue types, including the lung (I. Spasojevic et al.,Free Radic Biol Med 45: 943-949; B. Gauter-Fleckenstein et al., FreeRadic Biol Med 48: 1034-1043). Penile tissues had the lowest levels ofMnTE-2-PyP, but were still detectable. We conclude that adequate levelsof MnTE-2-PyP are able to reach the urogenital system when MnTE-2-PyP isadministered i.p.

A known side-effect of radiation exposure is weight loss. Animals wereweighed throughout the course of the experiment (FIG. 3). Irradiatedrats lost on average 23.9% more weight than non-irradiated rats.Irradiated rats injected with MnTE-2-PyP only lost 5.9% more weight thanMnTE-2-PyP injected, non-irradiated, control animals. The irradiatedrats receiving MnTE-2-PyP lost significantly less weight as compared toirradiated rats not receiving MnTE-2-PyP at all time points measured(FIG. 3).

Throughout the course of the experiment, phenotypic changes wereobserved between the treatment groups. Epilation was observed in thelower abdominal region in irradiated rats (data not shown); however,treatment with MnTE-2-PyP markedly blocked epilation in the radiationexposed area (data not shown). The skin of PBS injected irradiated ratshad significantly more erythema and moist desquamation as compared tothe skin of irradiated animals injected with MnTE-2-PyP (data notshown). An additional observation was a significant radiation-inducedatrophy of the rat testes at 12 weeks post-irradiation (data not shown).The testes were ˜40% the size of non-irradiated testes. However, we didnot observe a change in the testis size in irradiated rats receivingMnTE-2-PyP compared to control, suggesting that MnTE-2-PyP also protectsthe testis from radiation-induced atrophy/damage.

At 12 weeks post-radiation therapy, histological analysis was performedon the prostate and penile tissues. Representative images of hematoxylinand eosin staining within the prostate demonstrate that MnTE-2-PyPprotected the prostate epithelial glands from atrophy and loss ofprostatic epithelial architecture in the irradiated rats (data notshown). Moreover, penile tissues were stained using Masson's trichromestain to characterize tissue fibrosis. The trichrome staining revealedthat MnTE-2-PyP prevented radiation-induced loss of smooth muscle andaccumulation of collagen in penile tissues (data not shown), suggestingthat MnTE-2-PyP protects penile tissue from radiation-induced fibrosis.

Erectile dysfunction is a common side-effect associated with prostatecancer radiation therapy. In our radiation model, we observedsignificant reduction in the intracavernous pressure in rat penises 12weeks post-irradiation as compared to non-irradiated saline controls(FIG. 4). A reduction in intracavernous pressure is a direct measure oferectile dysfunction. Irradiated rats lost >50% of their intracavernouspressure; however, the irradiated rats receiving MnTE-2-PyP werecompletely protected from intracavernous pressure loss. Thus, MnTE-2-PyPserved to protect against radiation-induced erectile dysfunction.

The foregoing is illustrative of the present invention, and is not to beconstrued as limiting thereof. The invention is defined by the followingclaims, with equivalents of the claims to be included therein.

That which is claimed is:
 1. A method of treating or inhibiting thedevelopment of erectile dysfunction and/or incontinence following pelvicradiation treatment in a subject in need thereof, comprisingadministering to said subject an active agent in a treatment effectiveamount, wherein said active agent is a compound of Formula B1:

wherein: each R is C₁₋₁₂ alkyl; each A is an independently selectedhydrogen or an electron-withdrawing group; M is a metal selected fromthe group consisting of manganese, iron, copper, cobalt, nickel andzinc; and Z³¹ is a counterion; or a pharmaceutically acceptable saltthereof, and wherein said active agent is administered to said subjectin an amount from about 0.01 milligram per kilogram subject body weight(mg/kg) to about 10 mg/kg.
 2. The method of claim 1, wherein saidtreatment includes radiation treatment and said active agent isadministered to said subject: prior to the onset of radiation treatmentof said subject, during radiation treatment of said subject, and/orfollowing radiation treatment of said subject.
 3. The method of claim 1,wherein said radiation treatment comprises external beam radiation,radioactive seed implantation therapy (brachytherapy), or a combinationthereof.
 4. The method of claim 1, wherein said radiation treatment isfor prostate cancer, cervical cancer, colorectal cancer, bladder cancer,uterine cancer, or endometrial cancer in said subject.
 5. The method ofclaim 4, wherein said prostate cancer comprises Stage I, Stage IIA,Stage IIB, Stage III, or Stage IV prostate cancer.
 6. The method ofclaim 1, wherein said compound has the structure:

wherein Z³¹ is a counterion.